Current analgesics are limited due to adverse side effects including addiction and organ damage. Indeed, the need for novel analgesics is highlighted as a High Priority by NCCAM. Our long-term goal is to harness the potential of probiotics to develop new probiotic analgesics for treating pelvic pain of interstitial cystitis/bladder pain syndrome (IC). We previously examined the molecular basis that discriminates the symptomatic response to uropathogenic E. coli (UPEC) from the lack of response to E. coli clinically associated with asymptomatic bacteriuria (ASB). We have now accumulated compelling preliminary data demonstrating that E. coli-based probiotics alleviate both acute and chronic pelvic pain in clinically-relevant murine models that recapitulate key aspects of IC. We hypothesize that the analgesic activity of E. coli-based probiotics results from acting on three receptors involved in bladder pain signals, Toll- like receptor 4 (TLR4), transient receptor potential cation channel subfamily V member 1 (TRPV1), and chemokine C-C motif receptor 2 (CCR2). These mechanistic studies will define the analgesic ligands of probiotic E. coli and define the roles for each receptor in mediating pelvic pain in murine models of IC. These studies are highly innovative, and we are unaware of other groups studying analgesic probiotics for chronic pelvic pain. This project will thus result in a new mechanistic understanding of probiotic analgesics and provide critical pre-clinical data to drive these probiotics into clinical trials for IC.